Analysis of Apoptotic Pathways by Western Blot

Study the effect of your compound on levels of cleaved caspase-3 and full length and cleaved PARP

Horizon offers western blot analysis of cleaved caspase-3 and full length and cleaved PARP to study the induction of apoptosis. Activation of the extrinsic or intrinsic pathway leads to the cleavage of caspase-3 (or caspase-7), which leads to the commitment to apoptotic death, and is thus considered a reliable apoptosis marker. PARP is involved in DNA repair, differentiation and chromatin structure formation. During apoptosis PARP is cleaved by caspase-3 and this impacts DNA fragmentation in the cell.

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Offering

  • Up to 16 treatment conditions can be analysed, e.g. 4 cell lines and 4 treatments at 2 time-points (n=1)
  • Standard preparation of the samples for western blotting
  • Detection: Cleaved caspase-3 and full length and cleaved PARP* (with commercial antibodies)
  • Loading control: beta-actin
  • Deliverables: Summary report and western blot images**

*Additional biomarkers can be tested if required (i.e. other pathway-specific cleaved caspases or selected pathway-related apoptotic markers).
Please enquire about a custom study. **Quantitation can be included for an additional fee

Case Study: Effect of Staurosporine on Cleaved PARP and Caspase 3 Protein Levels

  • Parental and isogenic HCT116 colon cancer cell lines were treated with staurosporine (0, 0.1, 1 mM) for 24 h.
  • Cells were harvested and total protein extracts probed for both PARP (full length versus cleaved) and activated caspase 3 (western blotting)
  • Conclusions: Both cleaved PARP and cleaved caspase 3 levels increase in a dose-dependent manner. The abundance of the cleaved proteins varies in the isogenic cell lines compared with the parental cell lines (stronger in CHEK2-/- cells and  weaker in CHEK2-/- TP53-/- cells).

Download the complete application note about our apoptosis assays

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